Assessment of Oxidative Stress Biomarkers in Rat Blood

Detalhes bibliográficos
Autor(a) principal: Sinzato, Yuri K. [UNESP]
Data de Publicação: 2023
Outros Autores: Rodrigues, Thiago, Cruz, Larissa L. [UNESP], Barco, Vinícius S. [UNESP], Souza, Maysa R. [UNESP], Volpato, Gustavo T., Damasceno, Débora C. [UNESP]
Tipo de documento: Artigo
Idioma: eng
Título da fonte: Repositório Institucional da UNESP
Texto Completo: http://dx.doi.org/10.21769/BioProtoc.4626
http://hdl.handle.net/11449/248617
Resumo: Redox status assessments are time-consuming, require a large volume of samples and great reagent amounts, and are not adequately described for methodological reproducibility. Here, the objective was to standardize redox balance determination, based on previously described spectrophotometric tests in pregnant rats, to improve precision, time dispensed, and the volume of samples and reagents, while maintaining accuracy and adequate cost benefits. This protocol summarizes oxidative stress markers, which focus on spectrophotometric tests for the assessment of thiobarbituric acid–reactive substances, reduced thiol groups, and hydrogen peroxide, as well as the antioxidant activity of superoxide dismutase, glutathione peroxidase, and catalase in washed erythrocyte and serum samples from full-term pregnant rats. For non-pregnant rats and other species, it is necessary to standardize these determinations, especially the sample volume. All measurements were normalized by the estimated protein concentrations in each sample. To establish optimum conditions for the reproducibility of the proposed methods, we describe all changes made in each assay’s steps based on the reference method reassessed for the new standardizations. Furthermore, the calculations of the concentrations or activities of each marker are presented. Thus, we demonstrate that the analysis of serum samples is easier and faster, but it is impossible to detect catalase activity. Furthermore, the proposed methods can be applied for redox balance determination, especially using smaller reagent amounts and lower sample volumes in lesser time without losing accuracy, as is required in obtaining samples during rat pregnancy.
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spelling Assessment of Oxidative Stress Biomarkers in Rat BloodBiomarkerLipid peroxidationOxidative stressPregnancyRedox statusRedox status assessments are time-consuming, require a large volume of samples and great reagent amounts, and are not adequately described for methodological reproducibility. Here, the objective was to standardize redox balance determination, based on previously described spectrophotometric tests in pregnant rats, to improve precision, time dispensed, and the volume of samples and reagents, while maintaining accuracy and adequate cost benefits. This protocol summarizes oxidative stress markers, which focus on spectrophotometric tests for the assessment of thiobarbituric acid–reactive substances, reduced thiol groups, and hydrogen peroxide, as well as the antioxidant activity of superoxide dismutase, glutathione peroxidase, and catalase in washed erythrocyte and serum samples from full-term pregnant rats. For non-pregnant rats and other species, it is necessary to standardize these determinations, especially the sample volume. All measurements were normalized by the estimated protein concentrations in each sample. To establish optimum conditions for the reproducibility of the proposed methods, we describe all changes made in each assay’s steps based on the reference method reassessed for the new standardizations. Furthermore, the calculations of the concentrations or activities of each marker are presented. Thus, we demonstrate that the analysis of serum samples is easier and faster, but it is impossible to detect catalase activity. Furthermore, the proposed methods can be applied for redox balance determination, especially using smaller reagent amounts and lower sample volumes in lesser time without losing accuracy, as is required in obtaining samples during rat pregnancy.Universidade Estadual PaulistaFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Laboratory of Experimental Research on Gynecology and Obstetrics Botucatu Medical School São Paulo State University (Unesp), São Paulo StateCenter of Natural and Human Sciences (CCNH) Federal University of ABC (UFABC), São Paulo StateLaboratory of System Physiology and Reproductive Toxicology Institute of Biological and Health Sciences Federal University of Mato Grosso (UFMT), Mato Grosso StateLaboratory of Experimental Research on Gynecology and Obstetrics Botucatu Medical School São Paulo State University (Unesp), São Paulo StateFAPESP: 2016/25207-5Universidade Estadual Paulista (UNESP)Universidade Federal do ABC (UFABC)Federal University of Mato Grosso (UFMT)Sinzato, Yuri K. [UNESP]Rodrigues, ThiagoCruz, Larissa L. [UNESP]Barco, Vinícius S. [UNESP]Souza, Maysa R. [UNESP]Volpato, Gustavo T.Damasceno, Débora C. [UNESP]2023-07-29T13:48:56Z2023-07-29T13:48:56Z2023-03-05info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articlehttp://dx.doi.org/10.21769/BioProtoc.4626Bio-protocol, v. 13, n. 5, 2023.2331-8325http://hdl.handle.net/11449/24861710.21769/BioProtoc.46262-s2.0-85151483851Scopusreponame:Repositório Institucional da UNESPinstname:Universidade Estadual Paulista (UNESP)instacron:UNESPengBio-protocolinfo:eu-repo/semantics/openAccess2024-08-16T14:12:50Zoai:repositorio.unesp.br:11449/248617Repositório InstitucionalPUBhttp://repositorio.unesp.br/oai/requestopendoar:29462024-08-16T14:12:50Repositório Institucional da UNESP - Universidade Estadual Paulista (UNESP)false
dc.title.none.fl_str_mv Assessment of Oxidative Stress Biomarkers in Rat Blood
title Assessment of Oxidative Stress Biomarkers in Rat Blood
spellingShingle Assessment of Oxidative Stress Biomarkers in Rat Blood
Sinzato, Yuri K. [UNESP]
Biomarker
Lipid peroxidation
Oxidative stress
Pregnancy
Redox status
title_short Assessment of Oxidative Stress Biomarkers in Rat Blood
title_full Assessment of Oxidative Stress Biomarkers in Rat Blood
title_fullStr Assessment of Oxidative Stress Biomarkers in Rat Blood
title_full_unstemmed Assessment of Oxidative Stress Biomarkers in Rat Blood
title_sort Assessment of Oxidative Stress Biomarkers in Rat Blood
author Sinzato, Yuri K. [UNESP]
author_facet Sinzato, Yuri K. [UNESP]
Rodrigues, Thiago
Cruz, Larissa L. [UNESP]
Barco, Vinícius S. [UNESP]
Souza, Maysa R. [UNESP]
Volpato, Gustavo T.
Damasceno, Débora C. [UNESP]
author_role author
author2 Rodrigues, Thiago
Cruz, Larissa L. [UNESP]
Barco, Vinícius S. [UNESP]
Souza, Maysa R. [UNESP]
Volpato, Gustavo T.
Damasceno, Débora C. [UNESP]
author2_role author
author
author
author
author
author
dc.contributor.none.fl_str_mv Universidade Estadual Paulista (UNESP)
Universidade Federal do ABC (UFABC)
Federal University of Mato Grosso (UFMT)
dc.contributor.author.fl_str_mv Sinzato, Yuri K. [UNESP]
Rodrigues, Thiago
Cruz, Larissa L. [UNESP]
Barco, Vinícius S. [UNESP]
Souza, Maysa R. [UNESP]
Volpato, Gustavo T.
Damasceno, Débora C. [UNESP]
dc.subject.por.fl_str_mv Biomarker
Lipid peroxidation
Oxidative stress
Pregnancy
Redox status
topic Biomarker
Lipid peroxidation
Oxidative stress
Pregnancy
Redox status
description Redox status assessments are time-consuming, require a large volume of samples and great reagent amounts, and are not adequately described for methodological reproducibility. Here, the objective was to standardize redox balance determination, based on previously described spectrophotometric tests in pregnant rats, to improve precision, time dispensed, and the volume of samples and reagents, while maintaining accuracy and adequate cost benefits. This protocol summarizes oxidative stress markers, which focus on spectrophotometric tests for the assessment of thiobarbituric acid–reactive substances, reduced thiol groups, and hydrogen peroxide, as well as the antioxidant activity of superoxide dismutase, glutathione peroxidase, and catalase in washed erythrocyte and serum samples from full-term pregnant rats. For non-pregnant rats and other species, it is necessary to standardize these determinations, especially the sample volume. All measurements were normalized by the estimated protein concentrations in each sample. To establish optimum conditions for the reproducibility of the proposed methods, we describe all changes made in each assay’s steps based on the reference method reassessed for the new standardizations. Furthermore, the calculations of the concentrations or activities of each marker are presented. Thus, we demonstrate that the analysis of serum samples is easier and faster, but it is impossible to detect catalase activity. Furthermore, the proposed methods can be applied for redox balance determination, especially using smaller reagent amounts and lower sample volumes in lesser time without losing accuracy, as is required in obtaining samples during rat pregnancy.
publishDate 2023
dc.date.none.fl_str_mv 2023-07-29T13:48:56Z
2023-07-29T13:48:56Z
2023-03-05
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/article
format article
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://dx.doi.org/10.21769/BioProtoc.4626
Bio-protocol, v. 13, n. 5, 2023.
2331-8325
http://hdl.handle.net/11449/248617
10.21769/BioProtoc.4626
2-s2.0-85151483851
url http://dx.doi.org/10.21769/BioProtoc.4626
http://hdl.handle.net/11449/248617
identifier_str_mv Bio-protocol, v. 13, n. 5, 2023.
2331-8325
10.21769/BioProtoc.4626
2-s2.0-85151483851
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv Bio-protocol
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.source.none.fl_str_mv Scopus
reponame:Repositório Institucional da UNESP
instname:Universidade Estadual Paulista (UNESP)
instacron:UNESP
instname_str Universidade Estadual Paulista (UNESP)
instacron_str UNESP
institution UNESP
reponame_str Repositório Institucional da UNESP
collection Repositório Institucional da UNESP
repository.name.fl_str_mv Repositório Institucional da UNESP - Universidade Estadual Paulista (UNESP)
repository.mail.fl_str_mv
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